Difference between revisions of "Part:BBa K2255007:Design"

 
Line 7: Line 7:
  
 
===Design Notes===
 
===Design Notes===
−
Using the logiciel SignalP 4.1, we saw that the cleavage is made between the alanine and the glutamate.
 
−
T--Aix-Marseille--M13pIII-Sequencesignal.jpeg
 
  
−
In order to gain flexibility, which will help the enzyme to cleave the signal sequence, we add two glycine and one serine residue which we retrotranslate, with the codon biais of E. coli K12.
+
The signal sequence is crucial for the excretion of p3 in the periplasm.<ref name="Heilpern">Heilpern, A. J. & Waldor, M. K. pIIICTX, a predicted CTXphi minor coat protein, can expand the host range of coliphage fd to include Vibrio cholerae. J. Bacteriol. 185, 1037–1044 (2003).</ref>
  
−
The signal sequence and D1-D2 sequence are designed to make fusion protein, thus we choose to make them Freiburg assembly standard with Rfc25 prefix and sufix. This will be helpful in order to assemble our biobrick.  
+
As we remove it with our construction of M13 [http://2017.igem.org/Team:Aix-Marseille/M13_Design M13 Design], we must put another one. We choose to use the one coming from M13 as we use ''E. coli'' to produce our phage.  
  
 +
In order to be functional, the signal peptide must be cut down from the rest of the protein. Thus, we must add the cleavage site. Using the logiciel SignalP 4.1, we saw that the cleavage is made between the alanine and the glutamate.
 +
 +
[[File:T--Aix-Marseille--M13pIII-Sequencesignal.jpeg|400px|center]]
 +
 +
In order to gain flexibility, which will help the enzyme to cleave the signal sequence, we add two glycine and one serine residue which we retrotranslate, with the codon biais of E. coli K12.
  
 +
The signal sequence and D1-D2 sequence are designed to make fusion protein, thus we choose to make them Freiburg assembly standard with Rfc25 prefix and sufix. This will be helpful in order to assemble our biobrick.
  
 
===Source===
 
===Source===

Revision as of 16:29, 26 September 2017


Signal sequence of p3 from M13 (Rfc25)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

The signal sequence is crucial for the excretion of p3 in the periplasm.[1]

As we remove it with our construction of M13 [http://2017.igem.org/Team:Aix-Marseille/M13_Design M13 Design], we must put another one. We choose to use the one coming from M13 as we use E. coli to produce our phage.

In order to be functional, the signal peptide must be cut down from the rest of the protein. Thus, we must add the cleavage site. Using the logiciel SignalP 4.1, we saw that the cleavage is made between the alanine and the glutamate.

T--Aix-Marseille--M13pIII-Sequencesignal.jpeg

In order to gain flexibility, which will help the enzyme to cleave the signal sequence, we add two glycine and one serine residue which we retrotranslate, with the codon biais of E. coli K12.

The signal sequence and D1-D2 sequence are designed to make fusion protein, thus we choose to make them Freiburg assembly standard with Rfc25 prefix and sufix. This will be helpful in order to assemble our biobrick.

Source

This sequence came from the genome of M13.

References

  1. ↑ Heilpern, A. J. & Waldor, M. K. pIIICTX, a predicted CTXphi minor coat protein, can expand the host range of coliphage fd to include Vibrio cholerae. J. Bacteriol. 185, 1037–1044 (2003).